This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Specific Aims Obesity and type 2 diabetes are the leading causes of end-stage renal disease. Vascular abnormality and renal dysfunction are commonly associated with diabetes mellitus. Our goal is to elucidate the regulatory mechanism(s) and functional consequences of cytochrome P450 (CYP) epoxygenases in the development and progression of vascular and renal dysfunction related to diabetic metabolic syndrome. We and others have established that CYP epoxygenase metabolites, epoxyeicosatrienoic acids (EETs), are potent vasodilators and have anti-hypertensive properties. EETs also possess anti-inflammatory actions that could protect the kidney from injury in hypertension and diabetes mellitus. There is evidence that an inability to increase renal EET production contributes to the development of hypertension in high salt or high fat diet-treated rats. .Moreover, we found that a down regulation of renal epoxygenases was associated with attenuated acetylcholine-induced relaxation in prediabetic obese rats. However, it is unclear whether the CYP epoxygenase pathway is affected by or contributes to the development and progression of vascular and renal dysfunction in diabetic metabolic syndrome. Our preliminary studies have shown a significant decrease in renal endothelial CYP2C23, a predominant rat kidney epoxygenase, in Zucker diabetic fatty (ZDF) rats. Furthermore, this decrease in CYP2C23 was correlated with a reduction of functional PPARa protein in the ZDF kidneys. Interestingly, recent reports have suggested that CYP modulation by PPARa ligands may play an important role in the regulation of vascular and renal function. In view of these results, pilot studies were conducted in our laboratory demonstrating that an induction of CYP enzymes by fenofibrate, a PPARa agonist, was associated with a reduction of blood pressure and an attenuation of glomerular hypertrophy and renal fibrosis in ZDF rats. Based on these results, we hypothesize that an inappropriate regulation of CYP epoxygenases may contrubite to the development and progression of vascular and renal dysfunction, and that overproduction of EET will protect against vascular and renal injury in diabetic metabolic syndrome (Diagram). Specifically, we will address the following three aims: Aim 1. To test the hypothesis that renal CYP enzymes are inappropriately regulated in diabetic metabolic syndrome, which may involve PPARa receptor. 1. Determine the expression level and activity of CYP enzymes and PPARa regulation in diabetic rats. 2. Evaluate the role of PPARa in the CYP regulation by fenofibrate using PPARa knockout mice. 3. Delineate the transcriptional and/or post-transcriptional mechanisms responsible for CYP regulation. 4. Identify the additional transcription factors involved in the CYP regulation by assessing the DNA binding activities of NF-kB, AP-1, SP1 and c-Jun. Aim 2. To test the hypothesis that overproduction of EET will lower blood pressure and protect against renal injury in diabetic metabolic syndrome. 1. Assess the effect of fenofibrate and PPOH (an epoxygenase inhibitor) on blood pressure in diabetic rats. 2. Determine the effects of fenofibrate and PPOH on kidney damage by evaluating the changes in kidney structure and molecular markers including nephrin, podocin and kidney injury molecule-1 in diabetic rats. 3. Characterize the action of EET on glomerular cell proliferation and cell cycle regulatory proteins Aim 2. To test the hypothesis that overproduction of EET will lower blood pressure and protect against renal injury in diabetic metabolic syndrome. 1. Assess the effect of fenofibrate and PPOH (an epoxygenase inhibitor) on blood pressure in diabetic rats. 2. Determine the effects of fenofibrate and PPOH on kidney damage by evaluating the changes in kidney structure and molecular markers including nephrin, podocin and kidney injury molecule-1 in diabetic rats. 3. Characterize the action of EET on glomerular cell proliferation and cell cycle regulatory proteins Aim 3 To test the hypothesis that a down-regulati,on of EET formation may contribute to the development and progression of renal vascular dysfunction and that overproduction of EET will improve renal microvascular structure and function in diabetic metabolic syndrome. 1. Evaluate the contribution of CYP metabolites to the altered renal vascular structure and function during the development of diabetic metabolic syndrome. 2. Determine the effect of CYP2C gene transfer on renal vascular function in diabetic metabolic syndrome. 3. Characterize the role of PPARa in the regulation of vascular function using PPARa knock out mice. The results generated from the project will identify potential therapeutic targets and loci for the prevention and treatment of obesity and diabetes-related kidney disease in humans